Nitric Oxide Interaction with Red Blood Cells

描述（由申请人提供）：一氧化氮（NO）通过其合成和失活之间的平衡来实现其生物学功能。NO的生物合成受到高度调控并有充分的记录，而其失活则鲜为人知。NO失活的主要途径包括与氧合血红蛋白[HbFe (ll) O2]和各种自由基的反应。传统观点认为，一氧化氮被HbFe (ll) O2失活是不受调控的，因为一氧化氮被认为是自由而迅速地扩散到红细胞（RBC）膜上。最近的研究结果表明NO转运到红细胞是由膜骨架蛋白控制的。红细胞消耗NO的速率可以通过细胞骨架结合蛋白（如Band 3）扰乱细胞骨架网络来调节。在特定的。铁-亚硝基血红蛋白[HbFe (ll) NO]（-0.1%）的形成增加了NO消耗率。这种调节因子在生理和病理上都很重要，因为HbFe (ll) NO是在缺氧时形成的，并且在各种条件下都能在人体中检测到。因此，本应用程序的目的是研究亚硝基溴介导的调控的生化机制，并确定其生理/病理作用。据推测，处于“超级T”状态的HbFe (ll) NO与Band 3结合，并将其种群转变为二聚体形式，从而使细胞骨架网络松动。由于缺氧条件下肺内可产生HbFe (ll) NO，因此HbFe (ll) NO调控的NO消耗可能参与缺氧肺血管收缩。此外，根据初步数据，我们进一步假设HbFe (ll) NO可减弱NO介导的冠状动脉舒张。具体目标1将利用生物化学和生物物理技术探索细胞骨架和带3蛋白的状态，研究参与这种调节的机制。具体目标2将集中在使用分离的猪肺和冠状动脉微血管的这种调节的功能作用。总之，这些结果将提示临床相关性和潜在的干预措施。

DESCRIPTION (provided by applicant): Nitric oxide (NO) achieves its biological functions through a balance between its synthesis and inactivation. The biosynthesis of NO is highly regulated and well documented, whereas its inactivation is much less understood. The major pathways for NO inactivation include reactions with oxygenated hemoglobin [HbFe (ll) O2] and various free radicals. Conventional wisdom suggests that NO inactivation by HbFe (ll) O2 is not regulated, since NO is thought to diffuse freely and rapidly across the red blood cell (RBC) membrane. Recent findings have shown that NO transport into RBCs is controlled by the membrane skeleton proteins. The rate of NO consumption by RBCs can be modulated by perturbing the cytoskeleton network through cytoskeleton binding proteins such as Band 3. In particular. formation of iron-nitrosyl-hemoglobin [HbFe (ll) NO] (-0.1%) increased the NO consumption rate. This regulator is of physiological and pathological importance as HbFe (ll) NO is formed during hypoxia and has been detected in humans under various conditions. The purpose of this application is thus to investigate the biochemical mechanisms underlying the nitrosylHb-mediated regulations and to determine their physiological/pathological roles. It is hypothesized that HbFe (ll) NO in the "super T" state binds to Band 3 and shifts its population to the dimer form, which loosens the cytoskeleton network. Since HbFe (ll) NO may be produced in the lungs under hypoxia, the HbFe (ll) NO regulated NO consumption may participate in hypoxic pulmonary vasoconstriction. Moreover, based on preliminary data, it is further hypothesized that HbFe (ll) NO attenuates the NO-mediated coronary vasodilation. Specific aim 1 will investigate the mechanisms involved in this regulation using biochemical and biophysical techniques which probe the state of cytoskeleton and Band 3 protein. Specific aim 2 will focus on the functional roles of this regulation using isolated porcine pulmonary and coronary microvessels. Together, these results will suggest clinical relevance and potential interventions.